tetano
Editor, Senior Moderator
Sci Rep
. 2024 Mar 25;14(1):7017.
doi: 10.1038/s41598-024-54183-w. Evaluating and optimizing Acid-pH and Direct Lysis RNA extraction for SARS-CoV-2 RNA detection in whole saliva
Brayden LaBute[SUP] #[/SUP][SUP] 1 [/SUP], Jackie Fong[SUP] #[/SUP][SUP] 1 2 [/SUP], Farinaz Ziaee[SUP] 3 [/SUP], Robert Gombar[SUP] 1 [/SUP], Mathew Stover[SUP] 1 [/SUP], Terry Beaudin[SUP] 1 [/SUP], Maria Badalova[SUP] 1 [/SUP], Qiudi Geng[SUP] 4 [/SUP], Ryland Corchis-Scott[SUP] 4 [/SUP], Ana Podadera[SUP] 3 [/SUP], Kyle Lago[SUP] 2 [/SUP], ZhenHuan Xu[SUP] 5 [/SUP], Fievel Lim[SUP] 5 [/SUP], Felix Chiu[SUP] 5 [/SUP], Minghua Fu[SUP] 5 [/SUP], Xiaofeng Nie[SUP] 5 [/SUP], Yuanmin Wu[SUP] 5 [/SUP], Corrina Quan[SUP] 6 [/SUP], Caroline Hamm[SUP] 2 6 [/SUP], R Michael McKay[SUP] 4 [/SUP], Kenneth Ng[SUP] 3 2 [/SUP], Lisa A Porter[SUP] 7 8 [/SUP], Yufeng Tong[SUP] 9 10 [/SUP]
Affiliations
COVID-19 has been a global public health and economic challenge. Screening for the SARS-CoV-2 virus has been a key part of disease mitigation while the world continues to move forward, and lessons learned will benefit disease detection beyond COVID-19. Saliva specimen collection offers a less invasive, time- and cost-effective alternative to standard nasopharyngeal swabs. We optimized two different methods of saliva sample processing for RT-qPCR testing. Two methods were optimized to provide two cost-efficient ways to do testing for a minimum of four samples by pooling in a 2.0 mL tube and decrease the need for more highly trained personnel. Acid-pH-based RNA extraction method can be done without the need for expensive kits. Direct Lysis is a quick one-step reaction that can be applied quickly. Our optimized Acid-pH and Direct Lysis protocols are reliable and reproducible, detecting the beta-2 microglobulin (B2M) mRNA in saliva as an internal control from 97 to 96.7% of samples, respectively. The cycle threshold (Ct) values for B2M were significantly higher in the Direct Lysis protocol than in the Acid-pH protocol. The limit of detection for N1 gene was higher in Direct Lysis at ≤ 5 copies/μL than Acid-pH. Saliva samples collected over the course of several days from two COVID-positive individuals demonstrated Ct values for N1 that were consistently higher from Direct Lysis compared to Acid-pH. Collectively, this work supports that each of these techniques can be used to screen for SARS-CoV-2 in saliva for a cost-effective screening platform.
. 2024 Mar 25;14(1):7017.
doi: 10.1038/s41598-024-54183-w. Evaluating and optimizing Acid-pH and Direct Lysis RNA extraction for SARS-CoV-2 RNA detection in whole saliva
Brayden LaBute[SUP] #[/SUP][SUP] 1 [/SUP], Jackie Fong[SUP] #[/SUP][SUP] 1 2 [/SUP], Farinaz Ziaee[SUP] 3 [/SUP], Robert Gombar[SUP] 1 [/SUP], Mathew Stover[SUP] 1 [/SUP], Terry Beaudin[SUP] 1 [/SUP], Maria Badalova[SUP] 1 [/SUP], Qiudi Geng[SUP] 4 [/SUP], Ryland Corchis-Scott[SUP] 4 [/SUP], Ana Podadera[SUP] 3 [/SUP], Kyle Lago[SUP] 2 [/SUP], ZhenHuan Xu[SUP] 5 [/SUP], Fievel Lim[SUP] 5 [/SUP], Felix Chiu[SUP] 5 [/SUP], Minghua Fu[SUP] 5 [/SUP], Xiaofeng Nie[SUP] 5 [/SUP], Yuanmin Wu[SUP] 5 [/SUP], Corrina Quan[SUP] 6 [/SUP], Caroline Hamm[SUP] 2 6 [/SUP], R Michael McKay[SUP] 4 [/SUP], Kenneth Ng[SUP] 3 2 [/SUP], Lisa A Porter[SUP] 7 8 [/SUP], Yufeng Tong[SUP] 9 10 [/SUP]
Affiliations
- PMID: 38527999
- PMCID: PMC10963802
- DOI: 10.1038/s41598-024-54183-w
COVID-19 has been a global public health and economic challenge. Screening for the SARS-CoV-2 virus has been a key part of disease mitigation while the world continues to move forward, and lessons learned will benefit disease detection beyond COVID-19. Saliva specimen collection offers a less invasive, time- and cost-effective alternative to standard nasopharyngeal swabs. We optimized two different methods of saliva sample processing for RT-qPCR testing. Two methods were optimized to provide two cost-efficient ways to do testing for a minimum of four samples by pooling in a 2.0 mL tube and decrease the need for more highly trained personnel. Acid-pH-based RNA extraction method can be done without the need for expensive kits. Direct Lysis is a quick one-step reaction that can be applied quickly. Our optimized Acid-pH and Direct Lysis protocols are reliable and reproducible, detecting the beta-2 microglobulin (B2M) mRNA in saliva as an internal control from 97 to 96.7% of samples, respectively. The cycle threshold (Ct) values for B2M were significantly higher in the Direct Lysis protocol than in the Acid-pH protocol. The limit of detection for N1 gene was higher in Direct Lysis at ≤ 5 copies/μL than Acid-pH. Saliva samples collected over the course of several days from two COVID-positive individuals demonstrated Ct values for N1 that were consistently higher from Direct Lysis compared to Acid-pH. Collectively, this work supports that each of these techniques can be used to screen for SARS-CoV-2 in saliva for a cost-effective screening platform.